The Optimum Concentration of N-Methyl D-Aspartate to Induce Dorsal Root Ganglion Neuron Activation through the N-Methyl D-Aspartate Receptor Pathway: Creating a Neuron Model For the in-vitro Study of Pain

authors:

avatar Ristiawan M Laksono 1 , avatar Handono Kalim 2 , avatar Mohammad S Rohman 3 , avatar Nashi Widodo 4 , avatar Muhammad R Ahmad , avatar Willy Halim 5

Department of Anesthesiology and Intensive Therapy, Faculty of Medicine, Brawijaya University, Indonesia
Department of Internal Medicine, Faculty of Medicine, Brawijaya University, Indonesia
Department of Cardiology and Vascular Medicine, Faculty of Medicine, Brawijaya University, Indonesia
Department of Biology, Faculty of Mathematics and Natural Science, Brawijaya University, Indonesia
Departement of Medicine, Faculty of Medicine Brawijaya University, Indonesia
Warning: No corresponding author defined!
Journal of Cellular & Molecular Anesthesia: Vol.9, issue 1; e150299
article type: Original Articles
DOI: https://doi.org/10.22037/jcma.v9i1.42426

how to cite: Laksono R M, Kalim H, Rohman M S, Widodo N, Ahmad M R, et al. The Optimum Concentration of N-Methyl D-Aspartate to Induce Dorsal Root Ganglion Neuron Activation through the N-Methyl D-Aspartate Receptor Pathway: Creating a Neuron Model For the in-vitro Study of Pain. J Cell Mol Anesth. 2024;9(1):e150299. https://doi.org/10.22037/jcma.v9i1.42426.

Abstract

Background: In the in-vitro study on chronic pain, the N-methyl D-Aspartate receptor (NMDAR) activation in the dorsal root ganglion (DRG) neuron became one of the most important mechanisms to activate the chronic pain pathways. NMDAR activation can be induced using an NMDAR agonist. No guidelines explain the NMDA optimum concentration to induce DRG neuron activation through the NMDAR pathway. This study aims to find the optimum concentration of NMDA to induce DRG neuron activation through the NMDAR pathway. Materials and Methods: We treat DRG neuron culture derived from the F11 cell line with 10, 20, 40, 60, 80, and 100 ?M NMDA. Phosphorylated extracellular signal-regulated kinase (pERK), an activated neuron biomarker, is measured using an immunocytochemistry assay as a neuron activation biomarker. We validate the NMDA optimum concentration by measuring intracellular Ca2+ level, mitochondrial membrane potential (??m), and cytosolic adenosine triphosphate (ATP) in the activated neuron. Those parameters are the downstream process following NMDAR activation and are related to neuron activity. Statistical analysis was performed using the One-Way ANOVA test with ?=5%. Results: We found that NMDA 80 ?M significantly had the highest pERK intensity and showed the most optimum neuron activation. Validation tests show an increase in intracellular Ca2+ influx and ??m. NMDA 80 ?M also causes significant depletion in the cytosolic ATP concentration related to neuron activation. NMDA 80 ?M induces neuron activation by increasing pERK, Ca2+ influx, ??m, and cytosolic ATP depletion. Conclusion: NMDA 80 ?M is the optimum concentration to induce DRG neuron activation through the NMDA receptor pathway.